sc 365404 santa cruz biotechnology pmdm2  (Santa Cruz Biotechnology)

Journal: Journal of Cancer

Article Title: BRD7 Stabilizes P53 via Dephosphorylation of MDM2 to Inhibit Tumor Growth in Breast Cancer Harboring Wild-type P53

doi: 10.7150/jca.67447

Figure Lengend Snippet: Bromodomain of BRD7 is required for interaction of p53 and MDM2 as well as stabilization of p53. (A) Western blot analysis for BRD7, p53, phosphorylated AKT, total AKT (t-AKT), and phosphorylated MDM2 (pMDM2) in MCF-7 cells transfected with empty vector or WT BRD7 expressed plasmid in the existence of AKT activator SC79. (B) Western blot analysis for BRD7, p53, phosphorylated AKT, total AKT (t-AKT), phosphorylated MDM2 (pMDM2) and total MDM2 (tMDM2) in MCF-7 cells transfected with empty vector or WT BRD7, or BRD7 ∆brd expressed plasmids. brd, bromodomain. (C) The interaction between p53 and pMDM2, or tMDM2 in MCF-7 cells transfected with empty vector or WT BRD7, or BRD7∆brd expressed plasmid. IP, immunoprecipitation; IB, immunoblotting. (D) Overexpression of WT BRD7 but not BRD7∆brd inhibited the ubiquitination of p53 in MCF-7 cells. Cells were transfected with plasmid vector or WT BRD7, or BRD7∆brd expressed plasmid. Cell lysates were immunoprecipitated with the p53 antibody and then immunoblotted with the ubiquitin antibody.

Article Snippet: Following incubation in 5% nonfat milk for 2 h, the membrane was then incubated with primary antibody (polyclonal rabbit anti-BRD7 (1:500 dilution), anti-Bcl-2 (1:500 dilution), anti-β-Tubulin (1:500 dilution), and anti-Bax (1:500 dilution) from Proteintech, Wuhan, China; polyclone rabbit anti-p53 (1:500 dilution), anti-pMDM2 (1:1000 dilution), anti-total-PARP (1:1000 dilution), anti-c-PARP (1:500 dilution), anti-AKT (1:1000 dilution), anti-pAKT (1:1000 dilution), anti-Bak (1:1000 dilution), anti-Flag (1:1000 dilution), and anti-p21 (1:1000 dilution) from Cell Signaling Technology, Danvers, MA; monoclonal rabbit anti-total-MDM2 (1:1000 dilution) from Abcam, Cambridge, UK, and polyclonal rabbit anti-cyclin D1 (1:1000 dilution) and monoclonal mouse anti-GAPDH (1:3000 dilution) from Santa Cruz, Dallas, Texas, USA overnight at 4 °C.

Techniques: Western Blot, Transfection, Plasmid Preparation, Immunoprecipitation, Over Expression

Journal: Journal of Cancer

Article Title: BRD7 Stabilizes P53 via Dephosphorylation of MDM2 to Inhibit Tumor Growth in Breast Cancer Harboring Wild-type P53

doi: 10.7150/jca.67447

Figure Lengend Snippet: IHC detection of tumor samples from xenografted tumor in mice. Representative images of immunohistochemistry staining for BRD7, p53, phosphorylated MDM2 at ser166 (pMDM2) and p21, and the markers of proliferation and apoptosis, Ki67 and c-PARP in tumor tissues, and quantification of integral optical density (IOD). Data are expressed as mean ± s.d. *p<0.05, **p < 0.01, ***p < 0.001.

Article Snippet: Following incubation in 5% nonfat milk for 2 h, the membrane was then incubated with primary antibody (polyclonal rabbit anti-BRD7 (1:500 dilution), anti-Bcl-2 (1:500 dilution), anti-β-Tubulin (1:500 dilution), and anti-Bax (1:500 dilution) from Proteintech, Wuhan, China; polyclone rabbit anti-p53 (1:500 dilution), anti-pMDM2 (1:1000 dilution), anti-total-PARP (1:1000 dilution), anti-c-PARP (1:500 dilution), anti-AKT (1:1000 dilution), anti-pAKT (1:1000 dilution), anti-Bak (1:1000 dilution), anti-Flag (1:1000 dilution), and anti-p21 (1:1000 dilution) from Cell Signaling Technology, Danvers, MA; monoclonal rabbit anti-total-MDM2 (1:1000 dilution) from Abcam, Cambridge, UK, and polyclonal rabbit anti-cyclin D1 (1:1000 dilution) and monoclonal mouse anti-GAPDH (1:3000 dilution) from Santa Cruz, Dallas, Texas, USA overnight at 4 °C.

Techniques: Immunohistochemistry, Staining

Journal: Journal of Cancer

Article Title: BRD7 Stabilizes P53 via Dephosphorylation of MDM2 to Inhibit Tumor Growth in Breast Cancer Harboring Wild-type P53

doi: 10.7150/jca.67447

Figure Lengend Snippet: Bromodomain of BRD7 is required for interaction of p53 and MDM2 as well as stabilization of p53. (A) Western blot analysis for BRD7, p53, phosphorylated AKT, total AKT (t-AKT), and phosphorylated MDM2 (pMDM2) in MCF-7 cells transfected with empty vector or WT BRD7 expressed plasmid in the existence of AKT activator SC79. (B) Western blot analysis for BRD7, p53, phosphorylated AKT, total AKT (t-AKT), phosphorylated MDM2 (pMDM2) and total MDM2 (tMDM2) in MCF-7 cells transfected with empty vector or WT BRD7, or BRD7 ∆brd expressed plasmids. brd, bromodomain. (C) The interaction between p53 and pMDM2, or tMDM2 in MCF-7 cells transfected with empty vector or WT BRD7, or BRD7∆brd expressed plasmid. IP, immunoprecipitation; IB, immunoblotting. (D) Overexpression of WT BRD7 but not BRD7∆brd inhibited the ubiquitination of p53 in MCF-7 cells. Cells were transfected with plasmid vector or WT BRD7, or BRD7∆brd expressed plasmid. Cell lysates were immunoprecipitated with the p53 antibody and then immunoblotted with the ubiquitin antibody.

Article Snippet: Following incubation in 5% nonfat milk for 2 h, the membrane was then incubated with primary antibody (polyclonal rabbit anti-BRD7 (1:500 dilution), anti-Bcl-2 (1:500 dilution), anti-β-Tubulin (1:500 dilution), and anti-Bax (1:500 dilution) from Proteintech, Wuhan, China; polyclone rabbit anti-p53 (1:500 dilution), anti-pMDM2 (1:1000 dilution), anti-total-PARP (1:1000 dilution), anti-c-PARP (1:500 dilution), anti-AKT (1:1000 dilution), anti-pAKT (1:1000 dilution), anti-Bak (1:1000 dilution), anti-Flag (1:1000 dilution), and anti-p21 (1:1000 dilution) from Cell Signaling Technology, Danvers, MA; monoclonal rabbit anti-total-MDM2 (1:1000 dilution) from Abcam, Cambridge, UK, and polyclonal rabbit anti-cyclin D1 (1:1000 dilution) and monoclonal mouse anti-GAPDH (1:3000 dilution) from Santa Cruz, Dallas, Texas, USA overnight at 4 °C.

Techniques: Western Blot, Transfection, Plasmid Preparation, Immunoprecipitation, Over Expression

Journal: Journal of Cancer

Article Title: BRD7 Stabilizes P53 via Dephosphorylation of MDM2 to Inhibit Tumor Growth in Breast Cancer Harboring Wild-type P53

doi: 10.7150/jca.67447

Figure Lengend Snippet: IHC detection of tumor samples from xenografted tumor in mice. Representative images of immunohistochemistry staining for BRD7, p53, phosphorylated MDM2 at ser166 (pMDM2) and p21, and the markers of proliferation and apoptosis, Ki67 and c-PARP in tumor tissues, and quantification of integral optical density (IOD). Data are expressed as mean ± s.d. *p<0.05, **p < 0.01, ***p < 0.001.

Article Snippet: Following incubation in 5% nonfat milk for 2 h, the membrane was then incubated with primary antibody (polyclonal rabbit anti-BRD7 (1:500 dilution), anti-Bcl-2 (1:500 dilution), anti-β-Tubulin (1:500 dilution), and anti-Bax (1:500 dilution) from Proteintech, Wuhan, China; polyclone rabbit anti-p53 (1:500 dilution), anti-pMDM2 (1:1000 dilution), anti-total-PARP (1:1000 dilution), anti-c-PARP (1:500 dilution), anti-AKT (1:1000 dilution), anti-pAKT (1:1000 dilution), anti-Bak (1:1000 dilution), anti-Flag (1:1000 dilution), and anti-p21 (1:1000 dilution) from Cell Signaling Technology, Danvers, MA; monoclonal rabbit anti-total-MDM2 (1:1000 dilution) from Abcam, Cambridge, UK, and polyclonal rabbit anti-cyclin D1 (1:1000 dilution) and monoclonal mouse anti-GAPDH (1:3000 dilution) from Santa Cruz, Dallas, Texas, USA overnight at 4 °C.

Techniques: Immunohistochemistry, Staining

Journal: PLoS ONE

Article Title: Vapor of Volatile Oils from Litsea cubeba Seed Induces Apoptosis and Causes Cell Cycle Arrest in Lung Cancer Cells

doi: 10.1371/journal.pone.0047014

Figure Lengend Snippet: ( A ) Immunoblot analysis of pMdm2 Ser 166 , p21 and p53 was analyzed in control or VCO exposed A549 cells at indicated time periods. β-actin used as internal loading control. ( B ) Immunoblot showed p21 level in p53 siRNA or Sp1 siRNA or their control siRNA transfected A549 cells exposed with or without VCO. ( C ) ChIP assay demonstrated VCO exposure increases binding of p53 to its response element (RE1 and RE2) on p21 promoter. ( D ) Cyclin D1-p21 interaction was increased with increasing the time of VCO exposure, which was shown by co-immunoprecipitation study. ( E ) BrdU incorporation in control and VCO treated A549 cells were examined by florescence microscopy. ( F ) FACS analysis showed cell cycle arrest at G1 to S phase as indicated by increased percentage of G 0 /G 1 cells with the decrease of S and G 2 /M phase cells. Bar represents 20 µm.

Article Snippet: The primary antibodies for pAkt (Thr308; sc-135650), pAkt1/2/3 (Ser473; sc-7985-R), Akt 1/2/3 (sc-8312), pPDK1 (Ser241; sc-101775), Bcl-xL (sc-7195), pBad (Ser136; sc-7999), Bad (sc-7869), pMdm2 (Ser166; sc-293105), p53 (sc-6243), p21 (sc-756), cyclin D1 (sc-753), poly[ADP-ribosyl]-polymerase (PARP) (sc-7150), Cytochrome c (sc- 7159) and β-actin (sc-130657) were purchased from Santa Cruz Biotechnology Inc., California, USA and mTOR (#2983) was procured from Cell Signaling Technology Inc., Danvers, MA, USA.

Techniques: Western Blot, Transfection, Binding Assay, Immunoprecipitation, BrdU Incorporation Assay, Microscopy